Project Abstract: The cumulative exposure to atherogenic lipoprotein particles, as well as the cholesterol mass carried within them, are the primary causal factors for atherosclerosis. Since it is the cumulative exposure to atherogenic lipoprotein particles that mediates atherosclerosis, the patterns of change in these particles over the early adult life course are central to the development of atherosclerotic cardiovascular disease (ASCVD). To date, the trajectories of lipoprotein particle number (LPN) and their determinants are not known. There are greater than 200 distinct single nucleotide polymorphisms (SNPs) that are known to be associated with cholesterol and triglyceride levels. However, in aggregate these SNPs explain only 12% of the population variance in any of these parameters. Diet, physical activity, and obesity explain 10-30% of the variance in cholesterol. The environmental and genetic influence on lipid concentrations, as well as the inter-individual variation in the effects of these variables, suggest that non-sequence dependent changes in gene expression, e.g., epigenetic modifications like DNA methylation (DNAm), may play a significant role in modulating LPN and serum cholesterol levels and their consequences. If DNAm mediates some of the adverse LPN trajectories, it may serve as an early marker of vulnerability to adverse lifestyle factors in terms of lipid homeostasis. Our objective is to explore distinct trajectories of LPN across early adult life, the environmental and epigenetic mediators of these trajectories, and the associations between these trajectories and ASCVD. Due to the extensive phenotyping of epigenetic, anthropometric, and dietary and other lifestyle patterns across 34 years of follow-up, serial serum sample availability, subclinical atherosclerosis measures, and assessment of ASCVD events, the CARDIA study provides an unparalleled opportunity to understand the multifactorial and complex factors that determine LPN-associated ASCVD risk. We plan to measure LPN in longitudinal samples from CARDIA with NMR spectroscopy and describe the trajectories in LPN across early adult life and their associated lifestyle factors, anthropometric characteristics, and epigenetic modifications. We will also quantify the associations between LPN and subclinical atherosclerosis and ASCVD events. Understanding these key mediators of LPN trajectories may help clinicians target individuals at high risk for adverse lipid homeostasis and identify molecular targets for future therapies to reduce ASCVD risk through modulation of LPN.